Traditional means of treating cancer have been surgery, radiation, and chemotherapy. Recently, novel therapeutic strategies like hyperthermia and heat shock protein (HSP)-based cancer therapies have shown great potential. However, the mechanism by which HSPs elicit anti-tumor immunity is thus far incompletely understood. This application will address this gap in the knowledge base by elucidating the cellular activation mechanism induced by extracellular HSP70. The objective of this particular application, which is the next step towards attainment of our long-term goal, is to understand how extracellular HSP70 activates immune responses with special emphasis on mechanisms of anti-tumor immunity. The central hypothesis proposed in this application is that HSP70 found in the extracellular milieu or expressed on tumors act as both chaperone and cytokine, a chaperokine, activates cells of the innate immune system, resulting in the launching of non-specific killing mechanisms within the tumor microenvironment. The rationale for the proposed research is that, once understanding has been obtained regarding mechanisms of HSP70-induced cellular activation, it is expected that it will become possible to enhance HSP-based cancer therapies by targeting important signal transduction intermediates, either pharmacologically or through molecular biological strategies. We plan to test our central hypothesis and accomplish the overall objective of this application by pursuing the following 3 specific aims: 1) To study signal transduction pathways activated by extracellular HSP70, 2) To biochemically characterize and identify the HSP70 surface receptor and, 3) To study the biological consequence of extracellular HSP70 and its role in hyperthermia-induced tumor killing. Successful completion of these studies will allow fundamental new knowledge to be obtained that is expected to significantly advance the general field of heat shock protein research.